Tazy Coolando - Who on your mind Prod by - Eddy Calvert X ADOTHEGOD


Main Themes:

Accurate cell population identification and analysis in flow cytometry. The document focuses on gating strategies that ensure reliable and interpretable data by isolating target cell populations and excluding unwanted events like debris, dead cells, and doublets.
Importance of gating controls for data accuracy. The use of controls such as FMOs and viability dyes is emphasized to set accurate gates, minimize spillover effects, and ensure the analysis focuses on viable cells.
Application of flow cytometry in immunology research. The guide uses T cell analysis as a practical example, highlighting the use of specific markers like CD3, CD4, and CD8 for immunophenotyping.

Most Important Ideas/Facts:

Flow cytometry gating is crucial for:
Excluding unwanted events: "Gating is the process of setting criteria to identify cell populations of interest based on specific parameters in flow cytometry. Gating helps: Exclude unwanted events: Filters out dead cells, debris, and doublets."
Focusing on single cells: "Isolates single, viable cells for precise data analysis."
Identifying specific cell populations: "Differentiates based on fluorescence markers, size, and granularity."

Basic gating strategy involves:
FSC/SSC gating: Using Forward Scatter (FSC) for size and Side Scatter (SSC) for granularity to define the primary cell population and exclude debris.
Single-cell gating: Employing FSC-A vs. FSC-H and FSC-W vs. FSC-H plots to eliminate doublets.
Dead cell exclusion: Utilizing viability dyes like 7-AAD, Propidium Iodide (PI), or Live/Dead stains to remove dead cells from the analysis.
Population-specific gating: Using fluorescently labeled antibodies against specific markers, like CD3, CD4, and CD8 for T cells, to identify and analyze target cell populations.

Importance of controls:
Fluorescence Minus One (FMO) controls: "FMO controls are critical in multicolor flow cytometry to set accurate gates... Use FMO controls to: Determine where to set boundaries between positive and negative populations."
Positive and negative controls: Help to accurately identify cell populations and set appropriate gates.

Practical tips for effective gating:
Adjust gates for each experiment: As cell populations can vary between samples.
Run controls: To ensure accurate identification and gating of cell populations.
Use sequential gating: Starting with broad gating and moving towards more specific gating steps.
Optimize compensation: Especially in multicolor experiments, to correct for spectral overlap between fluorophores.

Key takeaway: Flow cytometry gating is a critical process that requires careful planning, execution, and use of controls to ensure the accurate identification and analysis of cell populations in various research applications, including immunology.

Flow Cytometry Gating_ A Comprehensive Guide

Key Ideas & Facts:

gRNA Design is Crucial: The process starts with designing a guide RNA (gRNA) that specifically targets the desired DNA sequence. Key considerations for gRNA design include target specificity, the presence of a PAM sequence ("NGG" for SpCas9), and guide sequence length. Online tools such as Benchling, CHOPCHOP, and CRISPOR can assist with this process.
"Ensure the gRNA sequence is highly specific to the target site to minimize off-target effects."
Preparing CRISPR Components: Once the gRNA is designed, it needs to be synthesized or cloned, along with the Cas9 enzyme. Options for preparation include ordering synthetic gRNA, cloning gRNA sequences into expression plasmids, and choosing the appropriate Cas9 delivery method (protein, mRNA, or plasmid DNA).

Delivering CRISPR into Cells: The next stage involves delivering the CRISPR components (gRNA and Cas9) into the target cells. Various transfection methods can be used, including lipid-mediated transfection, electroporation, and viral vectors. The choice of method depends on the specific cell type and experimental requirements.
Selecting Edited Cells: After transfection, researchers may need to select or enrich the population of cells that have been successfully edited. This can be achieved through antibiotic selection (for plasmid-based systems), fluorescence-activated cell sorting (FACS), or magnetic-activated cell sorting (MACS).

Verifying the Edits: It's essential to confirm that the CRISPR-Cas9 system has successfully edited the target gene. This involves screening techniques like PCR, restriction enzyme digestion, and sequencing (Sanger or next-generation sequencing).
Analyzing the Impact of Mutations: The final step is to analyze the effects of the CRISPR-induced mutations on gene function and cell phenotype. Techniques for mutation analysis include Western blotting to detect protein expression changes, quantitative PCR (qPCR) to measure mRNA levels, and functional assays specific to the gene's role.
Thorough Documentation: Meticulous documentation throughout the entire workflow is essential for reproducibility and validation of experimental results.

Conclusion: The Assay Genie article emphasizes the importance of a systematic and well-documented approach to CRISPR-Cas9 genome editing. Each step, from gRNA design to mutation analysis, is critical for achieving precise, reliable, and reproducible results.

CRISPR-Cas9 Genome Editing Workflow

auth3ntic.

CARH4RTt

A new geek rock song from the upcomping album 'Flat Earth Society'.

I Wanna Throw John in a Wormhole

LE FORUM de la FUTUROTIQUE - 1987
St Hiaire de Riez 85 avec la Radio RVM
Raconté en exclusivité par Francis Petit - Correspondant Ouest France

FORUM FUTUROTIQUE - 1987 raconté en exclusivité par Francis Petit - Correspondant Ouest France

神ノ木高校 Bassline部

https://youtu.be/bkGCeyzjEiA?si=3egJsXfm1QeSM2-z

ほっとりみっと

here's a musical adaptation of the serious knowledge, important questions, powerful affirmations & inspired motivations that Shadowless Forms breathed into Juice Aleem (MC, vocalist, shouter,  screamer,  writer, workshop facilitator & many other things : http://ninjatune.net/artist/juice-aleem) who in turn breathed all that into this great piece :) 

http://juice-aleeminati.tumblr.com/post/135123558708/the-truth-and-hip-hop-global-lies-asian-the

the music is based on a 15 (mostly sample-based) instrumental tracks
that I composed between 2000 & 2002 before gradually stopping my compulsive sampling of all my favorite records :)

This suite features tiny bits of music by John Surman, Gary Burton, the Stranglers, Keith Jarrett, Art Ensemble Of Chicago, Clifford Gilberto, Jack Dejohnette, Eric Dolphy, Weather Report, Charles Mingus, John Coltrane Quartet, Charlie Parker, Kim Duk Soo, Frank Zappa, King Crimson, Olivier Messiaen, Reggie Workman & Art Davis (from Coltrane's "Africa Brass"), Pat Metheny & more ... 

.. all sampled with much Love¹² & much Respect¹² :)

I started working on the last instrumental (starting at 8min 14sec)in  2002, this one is almost sample-free, I played the percussions, the beat (on a Boss DR-202 Dr. Groove that a friend had lent me for some time), the thumb-piano/kalimba, the guitar & the alto sax (towards the end), I added a bit of synth & I made the bass line using a couple of notes played by Jaco Pastorius :) 

DISCLAIMER : the audio mix of these sources should be considered as a derivative work
as allowed under the "fair use" rule of the copyright law (section 107 of the Copyright Act 1976),
it is noncommercial and transformative in nature, uses no more of the original than necessary,
and has no negative effect on the market for the original work.

No copyright is claimed in this musical experiment and to the extent that material may appear to be infringed, I assert that such alleged infringement is permissible under "fair use" principles in U.S. copyright laws.

However, if you believe material has been used in an unauthorized manner, please contact me ! :D

Peace¹² שָׁלוֹם ܫܠܡ سَلاَم + Myriads of Blessings¹²
+ Much Love¹² <3 to you
& your loved ones
& their loved ones
& their loved ones'
loved ones
etc ...

Juice Aleem - The Truth And Hip Hop : Global Lies Asian

Michel m'a dit, je cherche du temps d'antenne, je lui ai répondu banco ! sur le Web je peux faire... Covid oblige, les deux compères sans jamais s'être rencontrés, se topent la main à distance... La radio Arturnews est née. Avec Michel Cavard au micro et Philippe Raspail à la technique web. Un duo de choc avec maintenant plusieurs centaines d'inteviews déjà réalisées et disponibles sur cette plateforme et sur arturnews.com

Unleashing the power of discovery! 🚀
At Assay Genie, we provide cutting-edge research tools to fuel breakthroughs in life sciences. From high-quality ELISA kits to innovative bioresearch reagents, we empower scientists worldwide to explore, innovate, and transform research into results. Follow us for insights into the world of biotechnology, exciting updates, and the latest trends in scientific discovery! 🌍🔬

SO we are really good at making beats and rapping on them and these are some of the most elite song you every hear yout lufe yay leys go

One of the best musical sensations coming out of the DMV (D.C, Maryland & Virginia) area, Tazy Coolando brings a fresh dynamic approach to Hip Hop and R&B scene with a cross-genre styling made his own. Pushing boundaries through blurring lines between the traditional standards of the two, his clever wordplay, delivery, and melodic harmonies will captivate and enthrall audiences across a wide range of music lovers. Timothy Furr Jr. aka Tazy Coolando was born on October 23rd, 1994 in Landover, MD. His biggest inspirations in music come from artists such as Whitney Houston, Dean Martin, Frank Sinatra, Little Willie John, Marvin Gaye, Issac Hayes, Ray Charles, and more. Aside from Hip Hop and R&B, Tazy also pulls from his passion in soul and jazz. He attended the performing arts school of Thomas Pullen and also gained experience performing in the choir. Tazy developed his skills through song writing and monologues which eventually led him to begin creating his own music at the age of 8.

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